Investigate whether the signal transduction cascade that down-regulates DNA replication after exposure of cells to ionizing radiation is initiated in the nucleus or in the cytoplasm. In response to ionizing radiation exposure, cells in S-phase down-regulate DNA replication activity by reducing the frequency of replicon initiation. Although this process has been studied extensively over the past forty years, the molecular mechanism is unknown. We have recently provided evidence for the operation of a signal transduction pathway and of the activation of inhibitory factors acting in trans to regulate DNA replication in I irradiated cells(1-3). A complete understanding of the regulatory process requires identification of the target molecules that ultimately mediate the inhibition, identification of the site where the signal activating the signal transduction cascade is generated, and finally identification of the proteins involved in the signal transduction cascade. The site of initiation of regulatory signal transduction cascades induced in response to radiation exposure remains controversial. Experiments using intact, unperturbed cells are required to clarify this initiation site. Measurements of DNA replication offer a useful endpoint that can be used to quantitat the response. The Columbia Alpha Particle Microbeam offers unique opportunities to probe these issues.